Project Summary Promising progress has been made in several aspects of the project to develop single-molecule RNA sequencing chips, but technical setbacks have delayed progress and exhausted the resources currently available. The main challenges have been (1) lack of reproducibility of the ALD-deposited dielectric layer used in the the tunneling devices; (2) contamination of the polio virus RNA polymerase used to control translocation and (3) lack of on-site electrical testing at Wake Forest where the helium-ion milling is carried out. In each case, we have identified the source of the problems and believe we have good solutions available. Supplemental funding is sought for increased manpower at ASU so that chip development and testing can be completed in the coming year. In addition, funding is sought to implement a new coating process at ASU and to pay for chips made to the new design. Supplemental funding is sought for Thomas Jefferson University for polymerase purification and RNA library preparation. Funding is sought for Wake Forest for on-site electrical testing. The requested supplement will not change the scope of the parent grant.